1. Technical Field
The present invention relates to a microorganism capable of simultaneous co-fermentation of mixed sugars and a method for producing butanol using the same.
2. Description of the Related Art
Butanol is a chemical intermediate with a wide range of applications such as biofuels, and is thus considered very an useful chemical.
In the related art, a method for producing butanol, acetone and ethanol by fermenting sugars using Clostridium strains was utilized in the early 1900's. As prices for petroleum declined and butanol could be produced at low cost by an oxo process, biological methods for producing butanol are replaced by a method for producing butanol in a petrochemical way. However, due to various environmental problems including global warming and the like originating from the use of petroleum resources, there has been an increasing need for an environmentally friendly method for producing butanol through microbial fermentation with renewable sources.
However, in order to produce butanol on an industrial scale using microorganisms, it is necessary that cost for biomass to be utilized as raw materials by microorganisms is inexpensive and the biomass is a non-food resource. In practice, in the case of producing butanol using traditional starch-based resources, it is known that raw material costs account for 60% of production cost. This stems from rising crop prices and low fermentation yield of strains. Therefore, in order to produce biobutanol economically on an industrial scale, renewable, inexpensive and non-food resources can be considered as biomass. It is apparent that cellulosic biomass is the resource satisfying such conditions.
Cellulosic biomass is composed of cellulose having β-1,4 linked glucose units and hemicelluloses (arabinoxylan, galactomannan and xyloglucan) composed of various pentoses and hexoses. When cellulosic biomass is hydrolyzed, hexoses such as glucose, mannose, galactose, pentoses such as xylose, arabinose, and the like, and disaccharides, such as cellobiose are produced. Thereamong, xylose is known as the second most abundant saccharide after glucose present in cellulosic biomass. However, in the case of microorganisms, specifically Clostridium acetobutylicum ATCC824, it is known that metabolism of other sorts of sugars is repressed when glucose and other sorts of sugars are present simultaneously, which is referred to as carbon catabolite repression (CCR) (Ounine K, Petitdemange H, Raval G, Gay R. 1985. Appl Environ Microbiol 49:874-8). Such a CCR phenomenon inhibits complete fermentation of mixed sugars in a lignocellulosic hydrolysate and thus reduces fermentation yield, thereby reducing fermentation capabilities of the strain. For example, although Clostridium sp. AH-1 (FERM-P 6093 ATCC39045) can utilize arabinose and xylose, it preferentially utilizes glucose, and then arabinose and xylose. Accordingly, glucose is first consumed, and then arabinose and xylose are utilized after expressing genes required in utilization of arabinose and xylose. Thus, in the case of continuous fermentation of mixed sugars using Clostridium sp. AH-1 (FERM-P 6093 ATCC39045), there are problems in that not only are arabinose and xylose accumulated in a culture solution but it also takes several hours to express genes required for their utilization. Therefore, there is a need for microorganisms capable of producing butanol by simultaneously fermenting mixed sugars in a lignocellulosic hydrolysate without CCR.
With the recent development of metabolic engineering technology and complete genome sequencing of Clostridium acetobutylicum, continuous efforts have been focused on more effective production of butanol. Further, studies relating to engineering of metabolic pathways have been actively performed. For example, reports say that, when a catabolite control protein A (ccpA) gene of Clostridium acetobutylicum is deleted, CCR phenomenon is alleviated, thereby allowing simultaneous co-fermentation of glucose and xylose (Ren C, Gu Y, Hu S, Wu Y, Wang P, et al. 2010. Metabolic Engineering 12:446-54). However, in this case, the degree of co-fermentation of glucose and xylose is negligible and the capabilities of the strain are not sufficient in terms of applicability on an industrial scale. Further, reports say that, when a gene encoding enzyme II of the D-glucose phosphoenolpyruvate-dependent phosphotransferase system (PTS) of Clostridium acetobutylicum is deleted and xylose transferase, xylose isomerase and xylulose 5-phosphatase (xylose kinase) are expressed, CCR is alleviated, thereby allowing simultaneous co-fermentation of glucose and xylose to produce butanol (Xiao H, Gu Y, Ning Y, Yang Y, Mitchell W J, et al. 2011. Appl Environ Microbiol 77:7886-95). However, this process also has limits in terms of commercial applicability since only about 5 g/L of xylose can be simultaneously co-fermented (namely, simultaneous co-fermentation of xylose is low), and productivity (0.31 g/L/h) and yield (16% (wt/wt)) are very low.
Furthermore, a lignocellulosic hydrolysate produced by pretreatment of cellulosic biomass including woody biomass or grass type biomass such as wood, empty fruit bunch (EFB), corn stalk, rice straw, and the like (hereinafter referred to as “lignocellulosic biomass”) contains unknown substances which may cause side-reactions during pretreatment of lignocellulosic biomass by acids or bases during saccharification, thereby inhibiting growth of microorganisms. Accordingly, in order to effectively ferment mixed sugars simultaneously, genetic engineering for simultaneous co-fermentation of mixed sugars as well as microorganisms having tolerance against inhibitory substances should be developed at the same time. However, microorganisms having tolerance against inhibitory substances and capable of simultaneous co-fermentation of mixed sugars on a commercially applicable scale have not yet been developed.